| Short-term physiological mechanisms for seawater (SW) acclimation in tilapia (Oreochromis mossambicus) were examined. As part of this research, we developed a technique for adherence and fixation of dissociated tilapia gill cells on coverslips that we optimized for subsequent immunocytochemistry of these cells. We used this method to quantify tilapia mitochondria-rich cells (MRCs) and other gill cells (non-MRCs) within cell cycle phases using laser scanning cytometry (LSC), after the tilapia's exposure to 2/3 SW. Freshwater (FW)-acclimated tilapia's MRCs increased mitotic activity at 8 h following SW exposure, whereas, MRCs and non-MRCs accumulated in G2 phase of the cell cycle peaking at 72 h. We interpret mitotic changes as indicative of epithelial reorganization whereas G2 arrest is likely used as a means for conserving energy as the fish deals with the osmotic stress. We also used this novel method with LSC to quantify the time course of apoptosis during 2/3 SW acclimation of tilapia. Apoptosis in MRCs showed increased caspase 3/7 activity and DNA fragmentation, starting at 6 h of salinity stress and remained elevated for at least 5 d, a result we also measured by caspase 3/7 microplate-reader-based assays. In parallel to induction of apoptosis, MRCs also showed G2 arrest, and an increase in NKA abundance per MRC and MRC size. Overall, we interpret these data as evidence for a significant role of apoptosis in the reorganization of MRC populations that takes place during tilapia SW acclimation. We also measured plasma osmolytes, cortisol, and respiration in these fish following 2/3 SW exposure to examine possible correlations with cellular events. Cortisol increased rapidly by 3 h and remained elevated until 5 d. Plasma osmolality, [Na+], and [Cl-] were elevated at 6-8 h, but did not peak until 1 d following exposure. Oxygen consumption increased at 1 d SW exposure vs. FW, while ventilation frequency increased immediately following salinity stress. Overall, cortisol appears to mediate plasma solute concentration changes and increases in whole-animal metabolism and respiratory rates. These changes probably result from cellular restructuring and associated energetic costs as a consequence of the cellular stress response in tilapia during salinity stress. |
